LOCUS       V033655                 9497 bp    DNA     circular SYN 01-JAN-1980
DEFINITION  synthetic circular DNA.
ACCESSION   V033655
VERSION     V033655
KEYWORDS    .
SOURCE      .
  ORGANISM  .
            .
FEATURES             Location/Qualifiers
     LTR             234..414
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
                     /label="5' LTR (truncated)"
     misc_feature    458..583
                     /note="packaging signal of human immunodeficiency virus
                     type 1"
                     /label="HIV-1 Ψ"
     misc_feature    1076..1309
                     /note="The Rev response element (RRE) of HIV-1 allows for
                     Rev-dependent mRNA export from the nucleus to the
                     cytoplasm."
                     /label="RRE"
     CDS             1493..1537
                     /codon_start=1
                     /note="recognized by the 2H10 single-chain llama nanobody"
                     /product="antigenic peptide corresponding to amino acids
                     655 to 669 of the HIV envelope protein gp41 (Lutje Hulsik
                     et al., 2013)"
                     /transl_table=1
                     /translation="KNEQELLELDKWASL"
                     /label="gp41 peptide"
     misc_feature    1804..1921
                     /note="central polypurine tract and central termination
                     sequence of HIV-1"
                     /label="cPPT/CTS"
     promoter        2011..2214
                     /note="human cytomegalovirus (CMV) immediate early
                     promoter"
                     /label="CMV promoter"
     CDS             2317..3591
                     /label="Smad3(NM_016769)"
                     /note="Smad3(NM_016769)"
                     /gene="Smad3"
     promoter        3656..3867
                     /note="core&nbsp;promoter for human elongation factor
                     EF-1α"
                     /label="EF-1α core promoter"
     LTR             3880..4148
                     /note="truncated 5' long terminal repeat (LTR) from human
                     T-cell leukemia virus (HTLV) type 1"
                     /label="5' LTR (truncated)"
     regulatory      4177..4186
                     /note="vertebrate consensus sequence for strong initiation
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
                     /label="Kozak sequence"
     CDS             4207..4869
                     /codon_start=1
                     /product="green fluorescent protein 2 from Pontellina
                     plumata, also known as ppluGFP2 (Shagin et al., 2004)"
                     /transl_table=1
                     /translation="PAMEIECRITGTLNGVEFELVGGGEGTPKQGRMTNKMKSTKGALT
                     FSPYLLSHVMGYGFYHFGTYPSGYENPFLHAINNGGYTNTRIEKYEDGGVLHVSFSYRY
                     EAGRVIGDFKVVGTGFPEDSVIFTDKIIRSNATVEHLHPMGDNVLVGSFARTFSLRDGG
                     YYSFVVDSHMHFKSAIHPSILQNGGPMFAFRRVEELHSNTELGIVEYQHAFKTPIAFA"
                     /label="CopGFP"
     CDS             4939..4992
                     /codon_start=1
                     /note="Eukaryotic ribosomes fail to insert a peptide bond
                     between the Gly and Pro residues, yielding separate
                     polypeptides."
                     /product="2A peptide from&nbsp;Thosea asigna virus capsid
                     protein"
                     /protein_id=""
                     /transl_table=1
                     /translation="EGRGSLLTCGDVEENPGP"
                     /label="T2A"
     CDS             4993..5592
                     /codon_start=1
                     /gene="pac from Streptomyces alboniger"
                     /note="confers resistance to puromycin"
                     /product="puromycin N-acetyltransferase"
                     /transl_table=1
                     /translation="MTEYKPTVRLATRDDVPRAVRTLAAAFADYPATRHTVDPDRHIER
                     VTELQELFLTRVGLDIGKVWVADDGAAVAVWTTPESVEAGAVFAEIGPRMAELSGSRLA
                     AQQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKGLGSAVVLPGVEAAERAGVPAFLETS
                     APRNLPFYERLGFTVTADVEVPEGPRTWCMTRKPGA*"
                     /label="PuroR"
     misc_feature    5593..6180
                     /note="woodchuck hepatitis virus&nbsp;posttranscriptional
                     regulatory element"
                     /label="WPRE"
     LTR             6254..6487
                     /note="self-inactivating&nbsp;3' long terminal repeat (LTR)
                     from HIV-1"
                     /label="3' LTR (ΔU3)"
     polyA_signal    6559..6680
                     /note="SV40 polyadenylation signal"
                     /label="SV40 poly(A) signal"
     rep_origin      6699..6834
                     /note="SV40 origin of replication"
                     /label="SV40 ori"
     primer_bind     complement(6872..6888)
                     /note="common sequencing primer, one of multiple similar
                     variants"
                     /label="M13 rev"
     protein_bind    6896..6912
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="The lac repressor binds to the lac operator to
                     inhibit transcription in E. coli. This inhibition can be
                     relieved by adding lactose or
                     isopropyl-β-D-thiogalactopyranoside (IPTG)."
                     /label="lac operator"
     promoter        complement(6920..6950)
                     /note="promoter for the E. coli lac operon"
                     /label="lac promoter"
     protein_bind    6965..6986
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence
                     of cAMP."
                     /label="CAP binding site"
     rep_origin      complement(7274..7862)
                     /direction=LEFT
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
                     replication"
                     /label="ori"
     CDS             complement(8033..8893)
                     /codon_start=1
                     /gene="bla"
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /product="β-lactamase"
                     /transl_table=1
                     /translation="MSIQHFRVALIPFFAAFCLPVFA,HPETLVKVKDAEDQLGARVGY
                     IELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEY
                     SPVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDR
                     WEPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRS
                     ALPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGA
                     SLIKHW*"
                     /label="AmpR"
     promoter        complement(8894..8998)
                     /gene="bla"
                     /label="AmpR promoter"
     primer_bind     9472..9488
                     /note="common sequencing primer, one of multiple similar
                     variants"
                     /label="M13 fwd"
ORIGIN
        1 acgcgtgtag tcttatgcaa tactcttgta gtcttgcaac atggtaacga tgagttagca
       61 acatgcctta caaggagaga aaaagcaccg tgcatgccga ttggtggaag taaggtggta
      121 cgatcgtgcc ttattaggaa ggcaacagac gggtctgaca tggattggac gaaccactga
      181 attgccgcat tgcagagata ttgtatttaa gtgcctagct cgatacaata aacgggtctc
      241 tctggttaga ccagatctga gcctgggagc tctctggcta actagggaac ccactgctta
      301 agcctcaata aagcttgcct tgagtgcttc aagtagtgtg tgcccgtctg ttgtgtgact
      361 ctggtaacta gagatccctc agaccctttt agtcagtgtg gaaaatctct agcagtggcg
      421 cccgaacagg gacctgaaag cgaaagggaa accagagctc tctcgacgca ggactcggct
      481 tgctgaagcg cgcacggcaa gaggcgaggg gcggcgactg gtgagtacgc caaaaatttt
      541 gactagcgga ggctagaagg agagagatgg gtgcgagagc gtcagtatta agcgggggag
      601 aattagatcg cgatgggaaa aaattcggtt aaggccaggg ggaaagaaaa aatataaatt
      661 aaaacatata gtatgggcaa gcagggagct agaacgattc gcagttaatc ctggcctgtt
      721 agaaacatca gaaggctgta gacaaatact gggacagcta caaccatccc ttcagacagg
      781 atcagaagaa cttagatcat tatataatac agtagcaacc ctctattgtg tgcatcaaag
      841 gatagagata aaagacacca aggaagcttt agacaagata gaggaagagc aaaacaaaag
      901 taagaccacc gcacagcaag cggccactga tcttcagacc tggaggagga gatatgaggg
      961 acaattggag aagtgaatta tataaatata aagtagtaaa aattgaacca ttaggagtag
     1021 cacccaccaa ggcaaagaga agagtggtgc agagagaaaa aagagcagtg ggaataggag
     1081 ctttgttcct tgggttcttg ggagcagcag gaagcactat gggcgcagcc tcaatgacgc
     1141 tgacggtaca ggccagacaa ttattgtctg gtatagtgca gcagcagaac aatttgctga
     1201 gggctattga ggcgcaacag catctgttgc aactcacagt ctggggcatc aagcagctcc
     1261 aggcaagaat cctggctgtg gaaagatacc taaaggatca acagctcctg gggatttggg
     1321 gttgctctgg aaaactcatt tgcaccactg ctgtgccttg gaatgctagt tggagtaata
     1381 aatctctgga acagattgga atcacacgac ctggatggag tgggacagag aaattaacaa
     1441 ttacacaagc ttaatacact ccttaattga agaatcgcaa aaccagcaag aaaagaatga
     1501 acaagaatta ttggaattag ataaatgggc aagtttgtgg aattggttta acataacaaa
     1561 ttggctgtgg tatataaaat tattcataat gatagtagga ggcttggtag gtttaagaat
     1621 agtttttgct gtactttcta tagtgaatag agttaggcag ggatattcac cattatcgtt
     1681 tcagacccac ctcccaaccc cgaggggacc cgacaggccc gaaggaatag aagaagaagg
     1741 tggagagaga gacagagaca gatccattcg attagtgaac ggatctcgac ggtatcggtt
     1801 aacttttaaa agaaaagggg ggattggggg gtacagtgca ggggaaagaa tagtagacat
     1861 aatagcaaca gacatacaaa ctaaagaatt acaaaaacaa attacaaaaa ttcaaaattt
     1921 tatcgatact agtattatgc ccagtacatg accttatggg actttcctac ttggcagtac
     1981 atctacgtat tagtcatcgc tattaccatg gtgatgcggt tttggcagta catcaatggg
     2041 cgtggatagc ggtttgactc acggggattt ccaagtctcc accccattga cgtcaatggg
     2101 agtttgtttt ggcaccaaaa tcaacgggac tttccaaaat gtcgtaacaa ctccgcccca
     2161 ttgacgcaaa tgggcggtag gcgtgtacgg tgggaggttt atataagcag agctcgttta
     2221 gtgaaccgtc agatcgcctg gagacgccat ccacgctgtt ttgacctcca tagaagattc
     2281 tagagctagc ctgatacgaa ctcggaattc gccaccatgt cgtccatcct gcccttcacc
     2341 cccccgatcg tgaagcgcct gctgggttgg aagaagggcg agcagaacgg gcaggaggag
     2401 aagtggtgcg agaaggcggt caagagcttg gtgaagaagc tcaagaagac ggggcagttg
     2461 gacgagctgg agaaggccat caccacgcag aacgtgaaca ccaagtgcat taccatcccc
     2521 aggtcactgg atggtcggct gcaggtgtcc catcggaagg ggctccctca cgttatctac
     2581 tgccgcctgt ggcgatggcc cgacctgcac agccaccatg aattacgggc catggagctc
     2641 tgtgagtttg ccttcaacat gaagaaggat gaagtgtgtg taaatcctta ccactatcag
     2701 agagtagaga cgccagttct acctccagtg ttggtgccac gccacaccga gatcccggcc
     2761 gagttccccc cactggatga ctacagccat tccattcccg agaacactaa cttccctgct
     2821 ggcattgagc cccagagcaa tattccagaa accccacctc ctggctacct gagtgaagat
     2881 ggagaaacca gtgaccacca gatgaaccac agcatggacg caggttctcc aaacctctcc
     2941 ccgaatccga tgtccccagc acacaataac ttggacctac agccagtcac ctactgtgag
     3001 ccggccttct ggtgctccat ctcctactac gagctgaacc agcgagttgg ggagacattc
     3061 cacgcctcac agccatccat gacagtagat ggcttcactg acccctccaa ctcggagcgc
     3121 ttctgcctgg gcctactgtc caatgtcaac cggaatgcag ccgtggaact tacaaggcga
     3181 cacattggga gaggtgtgcg gctctactac atcggagggg aggtctttgc ggagtgcctc
     3241 agtgacagtg ctattttcgt ccagtctccc aactgcaacc agcgctatgg ctggcacccg
     3301 gccactgtct gcaagatccc accaggctgc aacctgaaga tcttcaacaa ccaggaattt
     3361 gctgccctcc tagctcagtc tgtcaaccag ggctttgagg ctgtctacca gctgacgcgc
     3421 atgtgcacca tccgtatgag cttcgtcaaa ggctggggag cagagtacag gagacagaca
     3481 gtgaccagca ccccctgctg gattgagcta cacctgaatg gacccttgca gtggcttgac
     3541 aaggtcctca cccagatggg ttccccgagc atccgctgtt ccagtgtgtc ttagggatcc
     3601 ctgcgatatc cagcgcggcc gcgaaggatc tgcgatcgct ccggtgcccg tcagtgggca
     3661 gagcgcacat cgcccacagt ccccgagaag ttggggggag gggtcggcaa ttgaacgggt
     3721 gcctagagaa ggtggcgcgg ggtaaactgg gaaagtgatg tcgtgtactg gctccgcctt
     3781 tttcccgagg gtgggggaga accgtatata agtgcagtag tcgccgtgaa cgttcttttt
     3841 cgcaacgggt ttgccgccag aacacagctg aagcttcgag gggctcgcat ctctccttca
     3901 cgcgcccgcc gccctacctg aggccgccat ccacgccggt tgagtcgcgt tctgccgcct
     3961 cccgcctgtg gtgcctcctg aactgcgtcc gccgtctagg taagtttaaa gctcaggtcg
     4021 agaccgggcc tttgtccggc gctcccttgg agcctaccta gactcagccg gctctccacg
     4081 ctttgcctga ccctgcttgc tcaactctac gtctttgttt cgttttctgt tctgcgccgt
     4141 tacagatcca agctgtgacc ggcgcctacg ctagacgcca ccatggagag cgacgagagc
     4201 ggcctgcccg ccatggagat cgagtgccgc atcaccggca ccctgaacgg cgtggagttc
     4261 gagctggtgg gcggcggaga gggcaccccc aagcagggcc gcatgaccaa caagatgaag
     4321 agcaccaaag gcgccctgac cttcagcccc tacctgctga gccacgtgat gggctacggc
     4381 ttctaccact tcggcaccta ccccagcggc tacgagaacc ccttcctgca cgccatcaac
     4441 aacggcggct acaccaacac ccgcatcgag aagtacgagg acggcggcgt gctgcacgtg
     4501 agcttcagct accgctacga ggccggccgc gtgatcggcg acttcaaggt ggtgggcacc
     4561 ggcttccccg aggacagcgt gatcttcacc gacaagatca tccgcagcaa cgccaccgtg
     4621 gagcacctgc accccatggg cgataacgtg ctggtgggca gcttcgcccg caccttcagc
     4681 ctgcgcgacg gcggctacta cagcttcgtg gtggacagcc acatgcactt caagagcgcc
     4741 atccacccca gcatcctgca gaacgggggc cccatgttcg ccttccgccg cgtggaggag
     4801 ctgcacagca acaccgagct gggcatcgtg gagtaccagc acgccttcaa gacccccatc
     4861 gccttcgcca gatcccgcgc tcagtcgtcc aattctgccg tggacggcac cgccggaccc
     4921 ggctccaccg gatctcgcga gggcagagga agtcttctaa catgcggtga cgtggaggag
     4981 aatcccggcc ctatgaccga gtacaagccc acggtgcgcc tcgccacccg cgacgacgtc
     5041 cccagggccg tacgcaccct cgccgccgcg ttcgccgact accccgccac gcgccacacc
     5101 gtcgatccgg accgccacat cgagcgggtc accgagctgc aagaactctt cctcacgcgc
     5161 gtcgggctcg acatcggcaa ggtgtgggtc gcggacgacg gcgccgcggt ggcggtctgg
     5221 accacgccgg agagcgtcga agcgggggcg gtgttcgccg agatcggccc gcgcatggcc
     5281 gagttgagcg gttcccggct ggccgcgcag caacagatgg aaggcctcct ggcgccgcac
     5341 cggcccaagg agcccgcgtg gttcctggcc accgtcggcg tctcgcccga ccaccagggc
     5401 aagggtctgg gcagcgccgt cgtgctcccc ggagtggagg cggccgagcg cgccggggtg
     5461 cccgccttcc tggagacctc cgcgccccgc aacctcccct tctacgagcg gctcggcttc
     5521 accgtcaccg ccgacgtcga ggtgcccgaa ggaccgcgca cctggtgcat gacccgcaag
     5581 cccggtgcct gaaatcaacc tctggattac aaaatttgtg aaagattgac tggtattctt
     5641 aactatgttg ctccttttac gctatgtgga tacgctgctt taatgccttt gtatcatgct
     5701 attgcttccc gtatggcttt cattttctcc tccttgtata aatcctggtt gctgtctctt
     5761 tatgaggagt tgtggcccgt tgtcaggcaa cgtggcgtgg tgtgcactgt gtttgctgac
     5821 gcaaccccca ctggttgggg cattgccacc acctgtcagc tcctttccgg gactttcgct
     5881 ttccccctcc ctattgccac ggcggaactc atcgccgcct gccttgcccg ctgctggaca
     5941 ggggctcggc tgttgggcac tgacaattcc gtggtgttgt cggggaagct gacgtccttt
     6001 ccatggctgc tcgcctgtgt tgccacctgg attctgcgcg ggacgtcctt ctgctacgtc
     6061 ccttcggccc tcaatccagc ggaccttcct tcccgcggcc tgctgccggc tctgcggcct
     6121 cttccgcgtc tccgccttcg ccctcagacg agtcggatct ccctttggcc gcctccccgc
     6181 ctggtacctt taagaccaat gacttacaag gcagctgtag atcttagcca ctttttaaaa
     6241 gaaaaggggg gactggaagg gctaattcac tcccaacgaa aataagatct gctttttgct
     6301 tgtactgggt ctctctggtt agaccagatc tgagcctggg agctctctgg ctaactaggg
     6361 aacccactgc ttaagcctca ataaagcttg ccttgagtgc ttcaagtagt gtgtgcccgt
     6421 ctgttgtgtg actctggtaa ctagagatcc ctcagaccct tttagtcagt gtggaaaatc
     6481 tctagcagta gtagttcatg tcatcttatt attcagtatt tataacttgc aaagaaatga
     6541 atatcagaga gtgagaggaa cttgtttatt gcagcttata atggttacaa ataaagcaat
     6601 agcatcacaa atttcacaaa taaagcattt ttttcactgc attctagttg tggtttgtcc
     6661 aaactcatca atgtatctta tcatgtctgg ctctagctat cccgccccta actccgccca
     6721 gttccgccca ttctccgccc catggctgac taattttttt tatttatgca gaggccgagg
     6781 ccgcctcggc ctctgagcta ttccagaagt agtgaggagg cttttttgga ggcctagact
     6841 tttgcagaga cggcccaaat tcgtaatcat ggtcatagct gtttcctgtg tgaaattgtt
     6901 atccgctcac aattccacac aacatacgag ccggaagcat aaagtgtaaa gcctggggtg
     6961 cctaatgagt gagctaactc acattaattg cgttgcgctc actgcccgct ttccagtcgg
     7021 gaaacctgtc gtgccagctg cattaatgaa tcggccaacg cgcggggaga ggcggtttgc
     7081 gtattgggcg ctcttccgct tcctcgctca ctgactcgct gcgctcggtc gttcggctgc
     7141 ggcgagcggt atcagctcac tcaaaggcgg taatacggtt atccacagaa tcaggggata
     7201 acgcaggaaa gaacatgtga gcaaaaggcc agcaaaaggc caggaaccgt aaaaaggccg
     7261 cgttgctggc gtttttccat aggctccgcc cccctgacga gcatcacaaa aatcgacgct
     7321 caagtcagag gtggcgaaac ccgacaggac tataaagata ccaggcgttt ccccctggaa
     7381 gctccctcgt gcgctctcct gttccgaccc tgccgcttac cggatacctg tccgcctttc
     7441 tcccttcggg aagcgtggcg ctttctcata gctcacgctg taggtatctc agttcggtgt
     7501 aggtcgttcg ctccaagctg ggctgtgtgc acgaaccccc cgttcagccc gaccgctgcg
     7561 ccttatccgg taactatcgt cttgagtcca acccggtaag acacgactta tcgccactgg
     7621 cagcagccac tggtaacagg attagcagag cgaggtatgt aggcggtgct acagagttct
     7681 tgaagtggtg gcctaactac ggctacacta gaaggacagt atttggtatc tgcgctctgc
     7741 tgaagccagt taccttcgga aaaagagttg gtagctcttg atccggcaaa caaaccaccg
     7801 ctggtagcgg tggttttttt gtttgcaagc agcagattac gcgcagaaaa aaaggatctc
     7861 aagaagatcc tttgatcttt tctacggggt ctgacgctca gtggaacgaa aactcacgtt
     7921 aagggatttt ggtcatgaga ttatcaaaaa ggatcttcac ctagatcctt ttaaattaaa
     7981 aatgaagttt taaatcaatc taaagtatat atgagtaaac ttggtctgac agttaccaat
     8041 gcttaatcag tgaggcacct atctcagcga tctgtctatt tcgttcatcc atagttgcct
     8101 gactccccgt cgtgtagata actacgatac gggagggctt accatctggc cccagtgctg
     8161 caatgatacc gcgagaccca cgctcaccgg ctccagattt atcagcaata aaccagccag
     8221 ccggaagggc cgagcgcaga agtggtcctg caactttatc cgcctccatc cagtctatta
     8281 attgttgccg ggaagctaga gtaagtagtt cgccagttaa tagtttgcgc aacgttgttg
     8341 ccattgctac aggcatcgtg gtgtcacgct cgtcgtttgg tatggcttca ttcagctccg
     8401 gttcccaacg atcaaggcga gttacatgat cccccatgtt gtgcaaaaaa gcggttagct
     8461 ccttcggtcc tccgatcgtt gtcagaagta agttggccgc agtgttatca ctcatggtta
     8521 tggcagcact gcataattct cttactgtca tgccatccgt aagatgcttt tctgtgactg
     8581 gtgagtactc aaccaagtca ttctgagaat agtgtatgcg gcgaccgagt tgctcttgcc
     8641 cggcgtcaat acgggataat accgcgccac atagcagaac tttaaaagtg ctcatcattg
     8701 gaaaacgttc ttcggggcga aaactctcaa ggatcttacc gctgttgaga tccagttcga
     8761 tgtaacccac tcgtgcaccc aactgatctt cagcatcttt tactttcacc agcgtttctg
     8821 ggtgagcaaa aacaggaagg caaaatgccg caaaaaaggg aataagggcg acacggaaat
     8881 gttgaatact catactcttc ctttttcaat attattgaag catttatcag ggttattgtc
     8941 tcatgagcgg atacatattt gaatgtattt agaaaaataa acaaataggg gttccgcgca
     9001 catttccccg aaaagtgcca cctgacgtct aagaaaccat tattatcatg acattaacct
     9061 ataaaaatag gcgtatcacg aggccctttc gtctcgcgcg tttcggtgat gacggtgaaa
     9121 acctctgaca catgcagctc ccggagacgg tcacagcttg tctgtaagcg gatgccggga
     9181 gcagacaagc ccgtcagggc gcgtcagcgg gtgttggcgg gtgtcggggc tggcttaact
     9241 atgcggcatc agagcagatt gtactgagag tgcaccatat gcggtgtgaa ataccgcaca
     9301 gatgcgtaag gagaaaatac cgcatcaggc gccattcgcc attcaggctg cgcaactgtt
     9361 gggaagggcg atcggtgcgg gcctcttcgc tattacgcca gctggcgaaa gggggatgtg
     9421 ctgcaaggcg attaagttgg gtaacgccag ggttttccca gtcacgacgt tgtaaaacga
     9481 cggccagtgc caagctg
//
